AD is a neurodegenerative disorder and the primary cause of dementia. Currently there is no cure for this devastating disorder. The pathology of AD is characterized by two distinctive hallmarks: the ?-amyloid plaques primarily comprised of a small protein, amyloid-? (A?)1-3, and the neurofibrillary tangles composed of the hyperphosphorylated tau protein. A? is produced via a serial cleavage of the amyloid-? precursor protein (APP) through ?- and ?-secretase. The aggregation-prone A?42 peptide is essential to AD pathology and more prevalent than other A? species in cerebral ?-amyloid plaques. Thus, we have focused on developing a compound that may preferentially decrease A?42 levels and be an effective therapeutic for AD. We discovered a class of ?-secretase modulators (GSMs), a group of small molecules that specifically modulate ?-secretase cleavage in APP and thus preferentially lower A?42 levels without altering cleavage of other ?-secretase substrates, e.g. Notch. Recently, we generated a novel class of pyridazine-based soluble GSMs (SGSMs), with strong potency in lowering A?42 levels and high aqueous solubility. So far our best lead compound is SGSM15606, which displays outstanding potency in lowering A?42 levels (IC50 = 7 nM in cell-based studies). It also has excellent pharmacokinetics and pharmacodynamics properties including excellent oral bioavailability, long terminal half-life and low clearance. SGSM15606 is being prepared for an Investigational New Drug application enabling for a Phase-IA AD clinical trial (single ascending dose). Now over a hundred analogues of SGSM15606 have been made and have yet to be tested as backups for the forthcoming clinical trials. We hypothesize that we will be able to characterize the effects of these molecules on AD pathology to identify an ideal compound more favorable than SGSM15606. Notably, our studies focused on disease pathology in preclinical studies will enhance the success of candidate molecules in potential clinical trials. Specifically, we will first use two-dimensional (2D) AD cell models to characterize and prioritize GSM15606 and analogs that preferentially decrease A?42 levels (Aim 1; Years 1-2). We then will study the prioritized compounds on ?- amyloid pathology and tau pathology using our novel 3D human neural cell cultures of AD (Aim 2; Years 2-3). Finally, we will identify the top compounds that can both acutely and chronically attenuate AD pathology using the well-characterized AD transgenic Tg2576 mice (Aim 3; Years 4-5). Collectively, the results of the proposed studies should not only identify the final and ultimate compound for an AD clinical trial, but also should enhance our understanding of the biology of ?-secretase and the pathogenesis of AD.